The goal of this project is to examine the expression and regulation of genes encoding gonadal polypeptide hormones that are involved in controlling mammalian reproduction. Four protein hormones that modulate the synthesis and secretion of the pituitary gonadotropins have been described. These are the hypothalamic peptide gonadotropin-releasing hormone (GnRH), the related gonadal hormones inhibin and activin, and the recently described ovarian hormone follistatin. Inhibin, activin, and follistatin regulate the secretion of follicle-stimulating hormone (FSH) from the pituitary. Because FSH plays a crucial role in modulating the gametogenic and steroidogenic functions of the gonads, these regulatory hormones are likely to be important for homeostasis in the reproductive system. We have isolated cDNA clones from the rat for the alpha beta(A), and beta(B) chains of inhibin and activin, and for follistatin. We have used these cDNAs to examine expression of the corresponding mRNAs in reproductive issues using in situ hybridization histochemistry, and have begun to explore the regulation of these genes during the rat estrous cycle and during pregnancy. We now propose to extend these studies in three directions. Firstly, we plan to explore the molecular mechanisms by which these genes are regulated. We will develop in vitro assays, such as primary rat granulosa cell cultures, to directly examine the effects of steroid and polypeptide hormones on inhibin and activin gene expression. Granulosa cell cultures will also be utilized for transfection studies in which inhibin and activin gene regulatory elements will be characterized. Secondly, we will generate transgenic mice expressing inhibin and activin fusion genes. The alpha and beta precursors will be expressed from heterologous promoters to examine the physiological consequences of their inappropriate expression in animals. The alpha and beta gene promoters will be used to express convenient marker genes, in order to study the tissue-specific and developmentally-regulated expression of these promoters in transgenic mice. Lastly, we will examine the biological significance of activin and follistatin expression in several novel tissues. The activin mRNA is found in diverse extra-gonadal tissues, which is intriguing in light of its similarity to the TGF-beta family of growth and differentiation regulators. We will investigate when during development the beta-chain genes are expressed in these issues, and determine whether the genes are regulated in a manner similar to that observed in the gonads. We will examine extra-gonadal expression of the rat follistatin gene, pursuing our recent observation that this gene is highly expressed in decidual issue early in pregnancy. We expect these experiments to enhance our understanding of the roles inhibin, activin, and follistatin play in regulating normal reproduction, and to provide a framework for examining how alterations in their activities might lead to reproductive disorders or dysfunction.